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H A Rotbart

Publications and source records attributed to H A Rotbart.

At least 55 records · Page 3Linked to original sources
PubMed1990

Persistent enterovirus infection in culture-negative meningoencephalitis: demonstration by enzymatic RNA amplification.

Chronic meningoencephalitis due to enterovirus infection can occur in patients with antibody deficiencies. A modified polymerase chain reaction technique demonstrated persistent echovirus 11 infection in such a patient, despite negative routine viral cultures and negative routine nucleic acid hybridization. Although the sequence of echovirus 11 has not yet been determined, genomic conservation among the enteroviruses is significant, permitting detection of echovirus 11 with a primer pair and probe derived from enterovirus serotypes that have been fully sequenced. This study provides the first definitive evidence for the persistence of enterovirus infection with negative viral cultures.

Agammaglobulinemia↗
PubMed1990

Detection of Listeria monocytogenes by using the polymerase chain reaction.

A method was developed for detection of Listeria monocytogenes by polymerase chain reaction amplification followed by agarose gel electrophoresis or dot blot analysis with a 32P-labeled internal probe. The technique identified 95 of 95 L. monocytogenes strains, 0 of 12 Listeria strains of other species, and 0 of 12 non-Listeria strains.

Bacteria↗
PubMed1990

Enzymatic RNA amplification of the enteroviruses.

Enteroviruses are among the most common causes of childhood infection. Current diagnostic techniques are often too slow and too insensitive to benefit the patient optimally. This report describes a modified polymerase chain reaction technique by which enteroviral RNA can be amplified, over a few hours, to a level detectable by agarose mini-gel electrophoresis or nucleic acid hybridization or both. Three oligomeric regions of great homology among the enteroviruses were identified and designated as a potential primer pair and probe. With this combination, all 11 of the enterovirus serotypes tested, representing the major subgroups of these pathogens, were successfully amplified and detected. The sensitivity and rapidity of this new assay speak to its potential clinical applicability in the diagnosis of enterovirus infections.

Base Sequence↗
PubMed1990

Human parvovirus infections.

The B19 strain of parvovirus causes several distinct and important clinical diseases in humans. Aplastic crisis in patients with chronic hemolytic anemia, persistent bone marrow depression in immunocompromised individuals, and hydrops fetalis all result from direct infection of hematopoietic cells by the virus. Erythema infectiosum, arthritis, and purpuric vasculitis are postinfectious manifestations of B19 parvovirus infection.

Female↗
PubMed1989

Evidence for a novel picornavirus in human dermatomyositis.

We prepared RNA probes from cloned segments of human and murine enteroviruses (EVs) for in situ hybridization of skeletal muscle biopsies from patients with dermatomyositis (DM), polymyositis, other inflammatory myopathies, and noninflammatory muscle diseases, and from normal control subjects. A probe derived from Theiler's murine encephalomyelitis virus (TMEV) detected viral RNA within mononuclear cells of the interstitial connective tissue in 3 of 5 patients with adult-onset DM. None of these patients showed positive hybridization to probes derived from human EVs (poliovirus type 1 and Coxsackie virus B3) applied to subjacent sections of the same biopsies. The remaining 2 adult DM patients, 4 patients with childhood-onset DM, and 24 non-DM patients did not react with either TMEV or human enterovirus probes. Histochemical stains for esterase and immunoperoxidase stains for Mac-1 antigen in the 3 DM patients who reacted positively revealed positive cells in the same distribution as, but in far greater number than, those positive by in situ hybridization. Immunoperoxidase staining for HLA-DR antigens revealed positive cells in the same distribution and number as were seen with the TMEV probe. We conclude that an EV-like agent, more closely related to TMEV than to human EVs, may be associated with DM and that this agent is probably localized within muscle macrophages that express class II major histocompatibility complex antigens.

Adult↗
PubMed1989

Analysis of enteric coagulase-negative staphylococci from neonates with necrotizing enterocolitis.

We investigated the possible role of coagulase-negative staphylococci in necrotizing enterocolitis. Stool coagulase-negative staphylococci isolated from neonates during an outbreak of rotavirus-associated necrotizing enterocolitis were analyzed by a variety of techniques, including speciation, semiquantitative culturing, antimicrobial susceptibility, plasmid profiling and cytotoxicity assays. No differences were observed between the coagulase-negative staphylococci of symptomatic babies and those of asymptomatic controls.

Bacteriological Techniques↗
PubMed1988

Intracellular detection of sense and antisense enteroviral RNA by in situ hybridization.

We prepared sense (S) and anti-sense (AS) 3H-labelled single-stranded RNA probes by polymerase-directed in vitro transcription of a coxsackievirus B3-derived cDNA fragment cloned in the PGEM2 plasmid vector. The probes detected, by in situ hybridization, both S and AS forms of viral RNA in the cytoplasm of coxsackievirus-infected tissue culture cells. More S (genomic and messenger) RNA was present than AS (negative and replicative intermediate) RNA. We confirmed and quantitated this observation with slot-blot hybridization of lysates of infected cells in which the ratio of detectable S to AS RNA was 40:1. The selective detection and localization of both forms of RNA in infected cells with sensitive and specific bi-directional probes advances the applicability of in situ hybridization to the study of viral pathogenesis.

Cell Line↗
PubMed1988

Development and application of RNA probes for the study of picornaviruses.

Single-stranded RNA probes were developed from cloned cDNA fragments derived from four picornaviruses; poliovirus type 1, coxsackievirus B3, ECHOvirus 9 and Theiler's murine encephalomyelitis virus. In comparative testing with a nick-translated cDNA probe, the RNA probe was found to be 10-100-fold more sensitive. Hybridization conditions were optimized for RNA probes to picornavirus targets. Longer hybridization times gave an improved signal, as did a temperature of 50 degrees C. Formamide concentration had little effect on signal strength. A crude transcription mix performed as well or better as a probe than did more purified preparations of RNA. Finally, a combination of the three probes derived from human picornaviruses detected all 16 serotypes of human enteroviruses tested. Used individually, the four probes detected different spectra of animal and human picornaviruses, shedding more light on the interrelationships among these pathogens.

DNA↗
PubMed1988

Neonatal rotavirus-associated necrotizing enterocolitis: case control study and prospective surveillance during an outbreak.

After the death of a premature infant from rotavirus-associated necrotizing enterocolitis, we instituted prospective surveillance for this disease in our neonatal intensive care unit. During the 4-month study period an additional six cases of necrotizing enterocolitis and eight cases of hemorrhagic gastroenteritis occurred. Rotavirus infection was documented in 11 of these 15 symptomatic infants, in comparison with only eight rotavirus infections in 147 asymptomatic or minimally symptomatic babies (P less than 0.0001). Stools from 110 nursery personnel tested during the outbreak did not contain rotavirus. However, 12 of 59 staff members had serum IgM antibody against rotavirus, suggesting recent infection. In a case-control study we compared babies with severe gastrointestinal illness with a control group randomly selected from asymptomatic babies in the nursery during the time of the outbreak. Univariate analysis found six categorical variables and nine continuous variables that were significantly associated with disease. Multivariate logistic regression analysis, however, found only birth weight (P less than 0.0001), rotavirus infection (P less than 0.0001), and age at time of first nonwater feeding (P less than 0.02) to be associated with gastrointestinal illness. This study provides further evidence for the role of infection in some cases of neonatal necrotizing enterocolitis and hemorrhagic gastroenteritis.

Colorado↗
PubMed1988

Nonisotopic oligomeric probes for the human enteroviruses.

Oligonucleotide probes were prepared from highly conserved regions of human enteroviral genomes. These reagents were labeled with either 32P or alkaline phosphatase and were successfully used in blot assays to detect a wide variety of human enteroviruses, proving the potential utility of oligomeric sequences as pan-enteroviral probes. The availability of nonisotopic probes will ultimately make a hybridization assay for enteroviruses easier, shorter, and more adaptable for routine diagnostic laboratories.

Base Sequence↗
PubMed1987

RNA target loss during solid phase hybridization of body fluids--a quantitative study.

Recent applications of nucleic acid hybridization to the diagnosis of viral infections are limited by the sensitivity of the assay system and, hence, by the amount of target viral nucleic acid in the specimen. In the case of single-stranded RNA viruses, RNase activity in body fluids poses a potential obstacle to optimizing sensitivity. We added radioactively labelled, purified, single-stranded enteroviral RNA to various body fluids and noted significant loss of this 'target' RNA during the course of routine hybridization procedures. We confirmed that RNase activity was responsible for much of the target loss. Further characterization of the RNase activity found it to be rapidly acting (less than 15 s), concentrated (equally active at 10-fold dilution of body fluid) and potent--resulting in RNA breakdown products too small to be retained by any of six commercially available membrane filters. The RNase was active only in solution and was effectively inhibited by treatment of the body fluid with inhibitors prior to contact of the fluid with RNA. In contrast, when RNA within virions was added to body fluids, it was largely retained by membrane filters during the hybridization procedure. We conclude that detection of single-stranded RNA viruses in body fluids will depend upon the quantity of intact virions in the specimen. RNase inhibitors should be immediately added to body fluid specimens after collection to minimize the loss of RNA which will occur if virions are disrupted in transport and handling of the specimen.

Amniotic Fluid↗
PubMed1987

Neonatal enterococcal sepsis: case-control study and description of an outbreak.

An increased incidence of neonatal sepsis caused by Group D streptococci, specifically enterococci (GDE), during a recent 6-month period prompted a 5-year review of enterococcal sepsis in our neonatal intensive care unit. Sixteen episodes occurred in 14 babies. GDE accounted for 8 of 19 (42%) episodes of neonatal bacteremia during the epidemic period vs. only 8 of 159 (5%) episodes during the remaining 4.5 years (endemic period) (P less than 0.001). Blood isolates were all identified as Streptococcus faecalis. A combination of three strain-typing methods successfully distinguished the epidemic organisms from endemic nursery strains of GDE and from "background" GDE flora in the hospital. Nursery isolates were all susceptible to ampicillin, intermediate or resistant to the aminoglycosides and variably resistant to the newer cephalosporins. There were no differences in antibiotic susceptibilities of the GDE or in characteristics of the patients that were unique to the epidemic. Neonates with GDE sepsis had a mean birth weight of 913 g, a mean gestational age of 27 weeks and a mean age of onset of sepsis of 8.5 weeks. Twenty-five controls, matched for birth weight and admission date, were identified. Significant differences (P less than 0.05) between cases and controls included: use of a nonumbilical central line (71 vs. 32%); days central line in place (26.5 vs. 6.5 days); and bowel resection (29 vs. 4%). This is the first reported outbreak of S. faecalis sepsis in neonates. GDE are important nosocomial pathogens that must be considered in late onset neonatal sepsis.

Cross Infection↗
PubMed1986

Noma in children with severe combined immunodeficiency.

Three Native American children with severe combined immunodeficiency developed noma, a necrotizing gingivostomatitis not previously reported in this country. The similarity between the clinical findings and those observed in monkeys with simian AIDS prompted us to evaluate our patients and their families for human retroviral infection. Antibodies to HTLV-I or HTLV-III/LAV proteins were not identified in patients nor in their family members. Standard bacterial and viral cultures similarly failed to identify a suspect pathogen.

Antibodies, Viral↗
PubMed1985

Factors affecting the detection of enteroviruses in cerebrospinal fluid with coxsackievirus B3 and poliovirus 1 cDNA probes.

Enteroviruses are common pathogens of meningitis and encephalitis, and infections are often difficult to distinguish clinically from bacterial and herpetic infections of the central nervous system. An array of enteroviruses added to cerebrospinal fluid in reconstruction experiments were detected by a dot hybridization assay. Optimal handling and processing conditions for infected cerebrospinal fluid were established, and the effect on the hybridization reaction of humoral and cellular components of the inflammatory response was determined. Six hybridization probes, derived from poliovirus 1 and coxsackievirus B3, were then tested, singly and in combinations, to optimize the sensitivity and spectrum of the assay. Implications for enteroviral taxonomy based on these experiments are discussed.

DNA↗
PubMed1985

Haemophilus influenzae type b septic arthritis in children: report of 23 cases.

Twenty-three cases of Haemophilus influenzae type b septic arthritis seen over a recent 5-year period are reviewed. The natural history of the disease includes a mean three days of fever and joint symptoms prior to hospitalization, often accompanied or immediately preceded by a viral illness and/or otitis media. Concurrent H influenzae type B meningitis was present in 30% of patients and concurrent osteomyelitis in 22%. Infants remained febrile in the hospital for a mean of 3.6 consecutive days. However, secondary and prolonged fevers were common. Clinical improvement in the joint examination was first seen at a mean of 2.5 days. Characteristic laboratory findings during recovery included a decline in total WBC count, neutrophil count, ESR, and hematocrit, with a concomitant increase in lymphocyte and platelet counts. Outpatient follow-up for a mean duration of 20 months found only two of 21 infants with residual impairment. The time to total healing in the remaining 19 infants, however, varied widely--from nine days to 17 months (mean of 4 months).

Anti-Bacterial Agents↗